Characterization and use of a novel AT-hook protein in Leishmania

描述(申请人提供)：利什曼病是一种使人衰弱的传染病，临床表现从皮肤损害到致命的内脏感染。这种由沙蝇传播的寄生虫病是由人类巨噬细胞感染属于利什曼原虫属的锥虫原虫引起的。目前利什曼病的治疗方法由于成本高、疗效低、毒性大以及耐药菌株的发病率增加而不足。因此，为利什曼病的治疗确定新的靶点至关重要。利用生物信息学策略，我们已经确定了一类以前在锥虫原生动物中没有研究过的利什曼原虫基因。这些基因编码AT-钩蛋白，这是一类参与多种细胞过程的蛋白质，包括DNA复制和转录。这些过程的重要性体现在它们对所有物种的保护上。DNA复制和转录一直是开发抗微生物药物的目标，包括针对艾滋病毒的抗逆转录病毒疗法，以减轻艾滋病患者的疾病负担。DNA复制和转录对于所有病原体的生存/复制至关重要，这使它们成为与利什曼原虫感染有关的疾病的诱人靶标。因此，阐明AT-钩蛋白在利什曼原虫中的作用有望为开发有效的治疗这种原虫寄生虫的新方法提供新的途径。顾名思义，AT-挂钩蛋白结合了富含AT的DNA。虽然利什曼原虫基因组富含GC，但它们含有富含AT的序列，其中一些位于与RNA转录启动相关的“链切换区”。为了了解AT-Hook蛋白在利什曼原虫生物学中的作用，我们选择了利什曼原虫的主要基因LmjF06.0270，该基因被预测编码一个180kD的AT-Hook蛋白。选择LmjF06.0270是因为LmjF06.0270在所有利什曼原虫物种中高度保守，但在其他真核生物中不存在，包括与之密切相关的锥虫，如锥虫和克里斯蒂亚。因此，LmjF06.0270是针对利什曼原虫所有物种引起的疾病的新靶点。在这一应用中，我们提出了表征LmjF06.0270分子功能的研究。此外，基于从考古细菌到后生真核生物保守的AT-钩蛋白的已知性质，我们建议利用LmjF06.0270来开发一种新的工具，该工具将显著加强所有利什曼原虫物种的遗传和分子研究。 公共卫生相关性：利什曼病威胁着全球3.5亿人，每年新增50万致命内脏利什曼病病例。世界卫生组织估计，这种经常使人虚弱和致命的疾病在全球88个国家广泛传播，影响到在流行地区工作的美国人，包括军事人员。这项研究的目标是开发治疗方法来减少这些数字，从而减轻痛苦，促进人类健康和生产力。

DESCRIPTION (provided by applicant): Leishmaniasis is a debilitating infectious disease, with clinical manifestations ranging from skin lesions to fatal visceral infections. This sandfly-borne parasitic disease results from infection of human macrophages with trypanosomatid protozoan parasites belonging to the genus Leishmania. Current treatments for leishmaniasis are inadequate due to their high cost, low efficacy, toxicity, and the increasing incidence of drug-resistant strains. Consequently, it is vitally important to identify new targets for therapies against Leishmania. Using bioinformatic strategies, we have identified a class of Leishmania genes previously not studied in trypanosomatid protozoans. These genes code for AT-hook proteins, a class of proteins that participate in multiple cellular processes, including DNA replication, and transcription. The importance of these processes is indicated by their conservation across all species. DNA replication and transcription have served as targets for the development of anti-microbials, including anti-retroviral therapies against HIV to reduce disease burden in AIDS patients. The essential nature of DNA replication and transcription for the survival/replication of all pathogens renders them attractive targets against diseases associated with Leishmania infections. Therefore, elucidating the role of AT-hook proteins in Leishmania promises new avenues through which effective novel therapeutics against this protozoan parasite may be developed. As their name suggests, AT-hook proteins bind AT-rich DNA. Although Leishmania genomes are highly GC-rich, they contain AT-rich sequences, some of which lie in "strand-switch regions" that are associated with initiation of RNA transcription. Toward understanding the role of AT-hook proteins in the biology of Leishmania, we have chosen to study the Leishmania major gene, LmjF06.0270, that is predicted to encode a 180 kD AT-hook protein. LmjF06.0270 was selected because it is highly conserved in all Leishmania species, but is absent in other eukaryote species, including closely-related trypanosomatids such as Trypanosoma and Crithidia. Therefore, LmjF06.0270 represents a novel target against diseases caused by all species of Leishmania. In this application, we have proposed studies to characterize the molecular functions of LmjF06.0270. In addition, based on known properties of AT-hook proteins conserved from Archaebacteria to metazoan eukaryotes, we propose to exploit LmjF06.0270 to develop a novel tool that will significantly augment genetic and molecular studies in all Leishmania species. PUBLIC HEALTH RELEVANCE: Leishmaniasis threatens 350 million people worldwide, with 500,000 new cases of fatal visceral leishmaniasis each year. The WHO estimates that this often debilitating and fatal disease is widespread in 88 countries worldwide, affecting Americans working in endemic areas, including military personnel. The goal of this research is to develop therapies to reduce these numbers, thus alleviating suffering and promote human health and productivity.